Kaposi's sarcoma-associated herpesvirus (KSHV), or human herpesvirus 8, is the causative agent of Kaposi's sarcoma (KS), which is a leading cancer in persons with AIDS, particularly in sub-Saharan Africa. The goals of this proposal are the study of the transmission and acquisition of KSHV using in vitro models of oral mucosa. Saliva is the only readily exchanged bodily fluid demonstrated to have infectious KSHV. This fact, and the epidemiology of KSHV infection, indicates saliva as the major infectious source. The source of infectious virus in the oral cavity will be investigated with the hypothesis that differentiation of the oral epithelium activates KSHV lytic replication. This will be carried out by: 1. Development and characterization of organotypic raft cultures derived from oral keratinocytes for the differentiation of in vitro epithelium. 2. Determination of KSHV replication in oral keratinocytes undergoing differentiation in organotypic raft cultures. 3. Determination of RTA promoter sequences controlling expression during epithelia development. 4. Because HIV infected individuals show higher rates and levels of KSHV shedding, the impact of HCMV and HIV on KSHV replication in in vitro epithelium will be examined. The infectious KSHV in saliva, and the presence of KSHV in groups without risk of sexually transmitted infections suggests the oral cavity as the site of acquisition, although little is known about the infection of oral tissue by KSHV. This proposal will investigate KSHV infection of oral mucosa with the following objectives: 1. To determine what cells of in vitro epithelium can be infected by KSHV, and can KSHV infect or egress from the apical and/or basolateral membranes of polarized keratinocytes. 2. To investigate the transmission of KSHV across the oral mucosa using in vitro organ culture models. 3. To determine the cells of oral mucosa organ cultures that are sites of primary KSHV infection.